Since the original descriptions of putrefaction by Hippocrates, sepsis has been recognized as a major cause of human suffering and mortality. Despite major advances made in understanding the systemic inflammatory response to infection, clinical trials of sepsis therapeutics have been repeatedly disappointing. These failure highlight the need for new, multidisciplinary perspectives into the onset, progression, and resolution of septic organ injury. The endothelial glycocalyx is a layer of glycosaminoglycans and associated proteoglycans lining the vascular surface. In vivo, the glycocalyx forms a substantial endothelial surface layer (ESL) that influences inflammation, endothelial permeability, and vascular tone-biologic processes highly relevant to sepsis pathophysiology. We have recently identified that the pulmonary ESL, by regulating exposure of endothelial surface adhesion molecules, serves a gatekeeping function controlling neutrophil transit into the lung. In response to an infectious stimulus, activated endothelial cells cleave the pulmonary ESL, allowing neutrophil adhesion and subsequent extravasation. Teleologically, this gatekeeping function would additionally require precise cellular control of ESL reconstitution, thereby limiting the magnitude of pulmonary inflammation. These processes of ESL repair, despite therapeutic relevance to patients with sepsis, have been unexplored. We hypothesize that a degraded pulmonary ESL is rapidly reconstituted in otherwise-healthy mice, allowing for maintenance of pulmonary vascular homeostasis. During sepsis, ESL reconstitution is delayed, contributing to the excessive pulmonary inflammation and edema characteristic of septic lung injury. Using state-of-the-art pulmonary intravital microscopy (E. Schmidt, Pulmonary/Critical Care, University of Colorado) and glycomic (R. Linhardt, Chemistry, Rensselaer Polytechnic Institute) approaches, we propose to (1) determine the mechanisms underlying pulmonary ESL reconstitution, (2) identify how these mechanisms are suppressed during sepsis, and (3) therapeutically manipulate these mechanisms to accelerate ESL reconstitution and attenuate septic lung injury. These multidisciplinary investigations, representing a highly novel collaboration within the field of sepsis, will be complemented by animal models of septic lung injury as well as analyses of biologic samples obtained from humans with severe sepsis. In summary, this proposal offers a new, multidisciplinary perspective on sepsis: that ESL integrity is a critical determinant of the onset and progression of septic organ injury. Investigating processes of ESL reconstitution may therefore identify novel therapeutic targets in a critical illness that, despite millennia of study, still lacks a clinically-efficacious, pathophysiology-targeted treatment.